Free fatty acids (FFAs) are vitally important components of lipids that modulate biological metabolism in various ways. Although the molecular structures are simple, the analysis of FFAs is still challenging due to their unique properties and wide concentration range. In the present study, a high-coverage liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established for the quantification of FFAs in serum samples using two structural analogues 5-(dimethylamino)naphthalene-1-sulfonyl piperazine (Dns-PP) and (diethylamino)naphthalene-1-sulfonyl piperazine (Dens-PP) as twin derivatization reagents. The Dns labeling of FFAs could significantly enhance their MS response via the introduction of the easily ionizable moiety of a tertiary amine-containing part and aid fragmentation in the multiple reaction monitoring (MRM) mode. Our results demonstrated that the detection sensitivities of FFAs were increased by 50–1500 fold compared with the nonderivatization method. At the same time, Dens-labeled standards were used as one-to-one internal standards to ensure accurate quantifications. Thirty-eight FFAs, covering short-, medium-, and long-chain, could be quantified in wide dynamic range with the lower limit of quantification (LLOQ) varied from 2 to 20 nM. Using this method, we analyzed serum FFAs in rat models of cisplatin-induced nephrotoxicity and irinotecan-induced gastrointestinal toxicity, respectively. The findings were further compared with those revealed by previous untargeted metabolomics. The results indicate that twin derivatization-based LC-MS provides a more accurate view of global FFA alternation and has great application potential in the fields of targeted metabolomics.

中文翻译：

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液相色谱-串联质谱法对游离脂肪酸进行高含量定量的双衍生化策略

游离脂肪酸（FFA）是脂质的重要组成部分，它们以多种方式调节生物代谢。尽管分子结构简单，但由于FFA的独特特性和宽浓度范围，其分析仍然具有挑战性。在本研究中，建立了一种高覆盖液相色谱-串联质谱（LC-MS / MS）方法，使用两种结构类似物5-（二甲基氨基）萘-1-磺酰基哌嗪（Dns）定量分析血清样品中的FFA。 -PP）和（二乙氨基）萘-1-磺酰基哌嗪（Dens-PP）作为双衍生试剂。FFA的Dns标记可以通过引入含有叔胺的部分的易于电离的部分来显着增强其MS反应，并在多反应监测（MRM）模式下帮助片段化。我们的结果表明，与非衍生化方法相比，FFA的检测灵敏度提高了50-1500倍。同时，使用Dens标记的标准品作为一对一的内部标准品，以确保准确定量。涵盖短链，中链和长链的38种FFA可以在宽动态范围内进行定量，其定量下限（LLOQ）在2到20 nM之间变化。使用这种方法，我们分别分析了顺铂诱导的肾毒性和伊立替康诱导的胃肠道毒性的大鼠模型中的血清FFA。将该结果与以前的非靶向代谢组学所揭示的结果进行了进一步的比较。